1. Field of the Invention
The present invention relates to a biologically pure culture of a microorganism belonging to Streptomyces sp. and capable of decomposing, or hydrolyzing, proteins recalcitrant to proteolysis, such as the perchloric acid-soluble protein (hereinafter referred to as “PSP”), to a method of producing a protease capable of decomposing proteins recalcitrant to proteolysis from the culture of the microorganism, to the isolated protease, to novel mutant strains of the microorganism, and to a method of treating materials containing proteins recalcitrant to proteolysis using the protease.
2. Description of the Prior Art
The technology of decomposing, or degrading, various proteins into peptides or amino acids is widely used in various industries, for example in producing preparations for medical use and food materials. The method of chemically decomposing proteins using hydrochloric acid or the like has excellent decomposition efficiency but may cause environmental pollution or the formation of undesirable byproducts due to severe decomposition conditions. Therefore, in human-related industries, in particular, methods of decomposition by means of proteases are utilized (cf. e.g. Japanese Patent Publication (JP Kokoku) H07-53106 and Laid-open Japanese Patent Application (JP Kokai) H11-75765). Thus, known in the art are compositions containing a Bacillus subtilis-derived enzyme which is thermostable in the middle to high temperature range and at the same time capable of depolymerizing proteins and can effectively decompose proteins normally recalcitrant to proteolysis (cf. e.g. Laid-open Japanese Patent Application (JP Kokai) 2001-037474), and proteolytic detergent compositions containing a Pyrococcus strain-derived superthermostable protease and excellent in detergency against proteinaceous stain components (cf. e.g. WO 00/61711).
The present inventors discovered a protein extractable from a hepatic cytoplasmic fraction with perchloric acid and having protein synthesis inhibiting activity, also referred to as “perchloric acid-soluble protein” or “PSP” (cf. e.g. J. Biol. Chem., 270, 30060, 1995). As a result of their continued study, they found that the inhibition of PSP expression results in cell proliferation. Further, they found that, when PSP is applied to proximal renal tubule cells, the intracellular expression of PSP is inhibited and, as a result, the proliferation of renal tubule cells is promoted, and thus PSP is effective in the treatment of nephropathies (cf. e.g. Laid-open Japanese Patent Application (JP Kokai) H11-292790). It is also known that PSP is structurally similar to those proteins called “abnormal prions” which cause BSE (bovine spongiform encephalopathy) and the like, is hardly decomposed and is preserved in various organisms, from animals to prokaryotes, and occurs universally in the environment (cf. e.g. Bioscience and Industry, 58 17-22, 2000).
Further, it has been reported that SAP, an extracellular alkaline serine protease, produced by Streptomyces sp. YSA-130 and homogeneously purified by CM-Sephadex column chromatography and crystallization, is a monomeric protein with a molecular weight of 19,000 (as determined by SDS-PAGE and gel filtration), that the amino acid composition and N-terminal sequence of SAP are very similar to those of other bacterial serine proteases such as Streptomyces griseus protease A and B, Lysobacter enzymogenes-derived α-soluble protease, and Nocardiopsis dassonvillei subsp. prasina OPC-210-derived alkaline serine protease NDP-I, that the optimum temperature and optimum pH for SAP are 60° C. and pH 11.5, respectively, and that SAP is stable at temperatures up to 50° C. and at pH 4 to 12, and that the activity of SAP is inhibited by Ag+, Hg+, Co2+, sodium dodecyl sulfate, N-bromosucciniride, diisopropyl fluorophosphate (DFP), 2,3-butanedione, 5,5′-dithiobis(2-nitrobenzoic acid) (DTNB), iodoacetic acid, N-ethylmaleimide (NEM), phenylmethanesulfonyl fluoride (PMSF) and phenylglyoxal (cf. e.g. Biosci. Biotech. Biochem., 58 (3), 470-474, 1994).
In recent years, a large number of microorganisms showing protease activity have been isolated from soils and biotic sludge. However, it has been considered that the utilization of these microorganisms cannot succeed in completely decomposing proteinaceous components, in particular proteinaceous components recalcitrant to proteolysis, contained in garbage, waste water, organic waste liquids, industrial wastes and the like. Accordingly, it is an object of the present invention to provide a novel microorganism high in protease activity and capable of decomposing proteins recalcitrant to proteolysis as contained in garbage, waste water, organic waste liquids, industrial wastes and the like, a protease produced by such microorganism and capable of decomposing proteins recalcitrant to proteolysis, and a method of utilizing the same.